The long term goals of the proposed research are to develop methods for efficient gene transfer into somatic cells as a means for treatment of human disease. Genetic diseases related to missing or altered gene products are good candidates for treaatment using these methods. Applications to cancer treatment include transfer of drug resistance genes into transplanted marrow allowing patients to tolerate higher doses of chemotherapy. It is generally agreed that retroviral gene transfer systems have the greatest potential for use in human somatic cell gene transfer. Bone marrow cells are ideal recipients for introduced genes because they are readily accessible, contain pluripotent self-renewing stem cells, and can be reliably reintroduced into animals. The aim of this investigation is to develop methods for efficient introduction of genetic material into bone marrow stem cells using retroviral vectors. Previous work by the Principal Investigator and others has shown that retroviral vectors allow highly efficient transduction of defined genes into cultured cells. Recently, retroviral gene transduction into bone marrow cells of intact mice has been demonstrated. We propose to extend these experiments using the canine model, which has been shown to be a good model for eventual human applications. A virus expressing a dominant-acting methotrexate-resistant dihydrofolate reductase gene will be used as a model virus. In comparison to genes previously inserted into retroviruses, this gene has the advantage that cells containing the gene can be selected in intact animals. The ability of the virus to confer methotrexate resistance to canine and human marrow in vitro, and to canine marrow cells transplanted into dogs will be tested. Methods of production of the model virus in the absence of replicating helper virus have been developed and will be more thoroughly tested in culture and in animals. Transfer of human disease-related genes will also be attempted in culture and in animals.